Oxidative stress is a major contributing factor in the pathogenesis of Parkinson's disease (PD). Mitochondria have long been implicated as the culprit for oxidative stress in PD. However, molecular sources for reactive oxygen species (ROS) in PD have not been clearly elucidated. A family of NADPH oxidase (NOX) is the first enzyme complex discovered which is specialized to generate superoxide. Here, we first demonstrate that the expression of Nox1, a Nox homologue, is increased in DA cells by oxidative stress such as 6-OHDA both in vivo and in vitro. Rac1, a key component of the Nox1 system, is also activated. Nox1 expression is increased in DA neurons of postmortem human brains from sporadic PD patients. Mutations in Leucine-rich-repeat-kinase 2 (LRRK2), the newly identified causative gene for PD also increase Nox1 expression and ROS generation in DA cells. Interestingly, Nox1 induction is affected by mitochondrial respiratory chain inhibitors, suggesting the interplay between mitochondrial dysfunction and Nox1 activation. Recent studies also suggest a prominent role of mitochondrial dysfunction in Nox1-mediated superoxide generation. Finally, inhibition of Nox1-mediated superoxide generation protects substantia nigra DA neurons from 6-OHDA- induced degeneration. These proposed studies will investigate 1) whether mitochondrial ROS plays a key role in Nox1 induction and activation, 2) the role of the Nox1/Rac1 system in degeneration of the nigrostriatal pathway by inhibition or activation of this system using AAV2-mediated gene transduction, and 3) whether LRRK2 mutations affect the activation of Nox1-mediated ROS production and consequential DA neurodegeneration. PUBLIC HEALTH RELEVANCE: Mitochondrial dysfunction and oxidative stress are strongly implicated in the pathogenesis of PD. The overall goal of this proposed study is to define the role of NADPH oxidase 1 and Rac1, the specialized superoxide generation system in degeneration of the dopaminergic nigrostriatal pathway. We will investigate whether mitochondria play a key role in the Nox1 induction and the intervention of the Nox system prevents DA neurodegeneration. Additionally, the study on the interaction between LRRK2 mutations and the Nox1/Rac1 activation may identify common molecular pathways involved in the pathogenesis of PD. Collectively, these results will help us to understand cellular mechanism governing the vulnerability of the DA nigrostriatal pathway to oxidative stress and lead to the development of novel therapeutic target.